AB0115 CD14+ MYELOID CELLS EXPRESS THE PDC MARKERS BDCA-2, BDCA-4, CD123 UPON DIFFERENTIATION IN BOTH HEALTHY INDIVIDUALS AND SLE PATIENTS
Authors: A. Psarras, Z. Wigston, E. Vital
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> Plasmacytoid dendritic cells (pDCs) are crucial mediators of the immune system linking innate and adaptive immune responses [1]. pDCs exhibit high expression of the IL-3 receptor (CD123) as well as other receptors such as BDCA-2 (CD303) and BDCA-4 (CD304). These molecules are used to identify pDCs in mechanistic studies and for therapeutic targeting. A recent clinical trial showed a reduction in interferon-stimulated gene expression and improvement in skin and joint inflammation in lupus patients who received treatment targeting BDCA-2 [2,3]. However, extensive functional and single-cell transcriptomic data have previously shown that pDCs are immunologically inert, not contributing to the type I IFN production seen in patients with systemic lupus erythematosus (SLE) [4,5]. <h3>Objectives</h3> To investigate whether the markers BDCA-2, BDCA-4, CD123 are expressed by myeloid cells other than pDCs. <h3>Methods</h3> Peripheral blood mononuclear cells (PBMCs) were obtained from healthy individuals and patients with SLE. PBMCs were analysed fresh or after 24-hour in vitro culture using flow cytometry for the pDC markers. CD14<sup>+</sup> monocytes were purified from freshly isolated PBMCs by negative selection. For M1 differentiation, monocytes were cultured for 6 days in the presence of GM-CSF; IFN-γ and LPS were added on day 6 and the cells were incubated for additional 24 hours. For M2 differentiation, monocytes were cultured for 6 days in the presence of M-CSF; IL-4 and IL-13 were added on day 6 and the cells were incubated for an additional 24 hours. After a 7-day culture, M1 and M2 cells were collected, stained for flow cytometry, and analysed using CytExpert (Beckman Coulter) software. <h3>Results</h3> On freshly analysed PBMCs, the expression of BDCA-2 and BDCA-4 was unique to pDCs in healthy controls. CD123 expression was also seen in a subset of CD14<sup>+</sup> monocytes as well as in CD3<sup>-</sup>CD19<sup>-</sup>CD14<sup>-</sup>CD11c<sup>-</sup>HLA-DR<sup>-</sup> cells. In some SLE patients, CD14<sup>+</sup> cells presented positive expression of BDCA-2, although at a lower level compared to pDCs (CD3<sup>-</sup>CD19<sup>-</sup>CD14<sup>-</sup>CD11c<sup>-</sup>HLA-DR<sup>+</sup>CD123<sup>high</sup>). After 24-hour in vitro culture, CD14<sup>+</sup> cells from both healthy and SLE patients presented strong expression of BDCA-2, BDCA-4, and CD123. After purification of CD14<sup>+</sup> cells and differentiation to either M1 (CD14<sup>+</sup>CD16<sup>+</sup>CD206<sup>+</sup>CD80<sup>high</sup>CD163<sup>low</sup>) or M2 (CD14<sup>+</sup>CD16<sup>+</sup>CD206<sup>+</sup>CD80<sup>low</sup>CD163<sup>high</sup>) phenotype, positive expression of these markers was seen. M1 macrophages showed high expression of BDCA-2 and BDCA-4 and almost universal expression of CD123. In contrast, M2 macrophages showed a meagre expression of BDCA-2 but universal expression of BDCA-4 and CD123. <h3>Conclusion</h3> We show that the BDCA-2, BDCA-4, CD123 markers -previously thought to be expressed exclusively by pDCs- are also expressed by differentiated CD14<sup>+</sup> myeloid cells in both healthy controls and patients with SLE. These findings can limit the significance of those markers to identify pDCs in tissue samples by simple immunohistochemistry. Finally, as monocytes have been shown to be the primary contributor to the interferon signature in blood, these data may explain how therapeutic targeting of BDCA-2 and related markers can impact disease activity and interferon activity in SLE patients. <h3>References</h3> [1]Psarras A, et al. Nat Rev Rheumatol. 2022 Oct;18(10):575-590. [2]Werth VP, et al. N Engl J Med. 2022 Jul 28;387(4):321-331. [3]Psarras, A. &amp; Vital, E.M. Anti-BDCA2 Antibody for Cutaneous Lupus Erythematosus. N Engl J Med 387, 1528-1529 (2022). [4]Psarras, A. et al. Nat Comm 11, 6149 (2020). [5]Billi, A.C. et al. Sci Transl Med 14, eabn2263 (2022). <h3>Acknowledgements:</h3> NIL. <h3>Disclosure of Interests</h3> Antonios Psarras: None declared, Zoe Wigston: None declared, Edward Vital Speakers bureau: AstraZeneca, Novartis, Paid instructor for: AstraZeneca, Novartis, Consultant of: UCB, Pfiser, Aurinia, Lilly, Roche, Genentech, Otsuka, GSK, Capella.